Research on animal, that is cloned animal, with separated embryo has been carried out these days. Recently, a sheep "Dolly" of the Roslin Institute in England is known as an example of cloned animal derived from somatic cell. In this example, it was confirmed that G.sub.0 phase synchronization of the cultured cells was very effective for synchronizing mitosis of cytoplasm of the recipient (embryo) and minimizing gene damage.
Further, research to find out a factor related to differentiation and maturation of various kinds of cell or inhibitory factor thereof has been carried out. For the above, it is necessary to control cell cycle of the cells into G.sub.0 phase. G.sub.0 phase synchronized cells have advantage of (1) responding clearly to growth factor and/or proliferation factor when the the cells are used as a model for the mechanism of cell in a living body, (2) minimizing gene damage when the cells are used as an donor of gene and (3) making their synchronization easy under the environments around the implanted cells.
Conventionally, there are serum restriction method or contact inhibition method as a method of synchronizing cells into G.sub.0 phase. In serum restriction method, human normal diploid cells are cultured to confluence in a growth medium (ES medium, 10% bovine fetal serum), followed by changing the medium to serum restricted medium (ES medium, 0.5% bovine fetal serum) and keeping the cells cultured by replacing the medium into fresh one on day 2 and 4, and then replacing it into growth medium (10% serum) on day 6 so as to stimulate proliferation thereof Usually, it takes about 20 hours to return to the progress of cell cycle by the stimulation of proliferation thereof. However, time to return to the progress of cell cycle by serum concentration, serum restricted time and stimulation with serum is different with respect to characters of cell and, therefore, it is necessary to adjust the conditions to each cell type. On the other hand, in contact inhibition method, when normal adhesive cells are cultured in a dish or a flask for cell culture, cells grow until colonies contact each other or colonies contact the wall of the dish or the flask (confluent state) and mitosis stops. Many cells at that time are considered to be in G.sub.0 phase of cell cycle. For example, cancer cell is considered to lose capability of contact inhibition and known to keep growing until colonies pile each other. However, in the case of bovine mammary gland epithelial cell, because G.sub.0 phase synchronization was incomplete using only contact inhibition method, the other method to solve the above problem is expected to develop.